Neuronal depolarization and neurotransmitter release underlie some of the most fundamental components of normal physiology and the etiology of brain pathophysiology. There is a tremendous need for high temporal resolution measurements of neurotransmitter release and its modulation of brain neuronal networks. While there has been progress in measuring neuronal depolarization in vivo in small animals, the current overall methodology of deployment, excitation and measurement of signal from voltage sensitive dyes (VSDs) commonly entails craniotomy and other invasive measures, and thus is currently only practical in rodent studies. We aim to develop a transformative brain imaging technique which will allow minimally invasive/non-invasive imaging of neuronal depolarization and related neurotransmitter release ultimately in the living human brain. While challenging methodologically, we believe that our team of multidisciplinary experts consisting of neuroscientists, neuropharmacologists, electrical and bioengineers, and brain imaging physicists and chemists, will be able to plan over a period of three years a practical and clear path to the development of such a potentially paradigm-shifting imaging technique. To do so, we propose three Aims. Aim 1 is to develop voltage sensitive probes for sub-millisecond measurements of membrane potentials and action potentials of cortical neurons in humans and other primates in vivo. Aim 2 will be to quantify highly temporally resolved neurotransmitter action with measures of lactate, pH, and redox potential changes in vivo. Finally, Aim 3 will pursue a pilot study of photoacoustic detection of neurotransmitter action by delivery of nanosecond pulses to intact skin and skull in response to changed absorption spectra of voltage or pH sensitive dyes. We hypothesize that we can also derive from these voltage depolarizations, regionally active neurotransmitter release, and through pharmacologic manipulation, help derive where the depolarizations have been modulated by neurotransmitters. This will allow understanding of depolarization waves that up to now have not been linked with neuropharmacology directly. Our approaches will be tested in the rodent brain and then translated into non-human primate brain. By the end of three years, we anticipate providing the evidence that it is feasible to carry out neurotransmitter modulation of neuroactivity, including neuronal depolarization, and to have developed a plan for building a brain imaging instrument to capture these events, enabling minimally-invasive procedures for transformative imaging of the human brain in health and disease.